IgA nephropathy (IgAN) is the most common primary glomerulonephritis and an important cause of end-stage kidney failure. It is a mesangioproliferative glomerulonephritis defined by IgA1 mesangial deposits. Although it has been speculated for some time that the pathogenesis of IgAN is driven by the deposition of circulating immune complexes, this has been difficult to prove due to the lack of animal models of IgAN. We have now developed and validated new protocols that permit the formation of engineered immune complexes in vitro together with a passive murine model of IgAN. These innovations together with our development of sensitive methods of analysis and extensive collaborations that permit testing of clinical correlations using well- characterized clinical cohots provide an unprecedented opportunity to elucidate the pathophysiology of IgAN. In IgAN, a fraction of IgA1 has galactose-deficient O-glycans (Gd-IgA1) that is present in circulating immune complexes with glycan-specific autoantibodies. We have defined the subsets of Gd-IgA1 and the anti-Gd-IgA1 autoantibodies that are present in these immune complexes and have used proteomic analysis to define the serum factors that associate with these complexes and may contribute to their pathogenic effects. Global kinase-activity profiling using an innovative peptide substrate microarray platform of human mesangial cells stimulated with Gd-IgA1 immune complexes identified robust tyrosine kinase activity, suggesting that the processes induced by these immune complexes in mesangial cells involve protein-tyrosine kinase signaling. Testing of protein-kinase inhibitors indicated that one of the inhibitors completely blocked immune complex- mediated mesangial cell proliferation in vitro, as well as in vivo in our new passive murine model of IgAN. Based on these novel results, we propose the hypothesis that Gd-IgA1-containing immune complexes represent a key hit in the pathogenesis of IgAN by activating mesangial cells through specific signaling pathways. A corollary to this hypothesis is that this immune-complex-driven signaling in mesangial cells can be blocked by small-molecular-mass inhibitors of protein kinases and thus represents a new therapeutic target(s). We will: 1) Define the characteristics of the Gd-IgA1-containing immune complexes from sera of patients with IgAN that activate human mesangial cells; 2) Determine the signaling pathways activated by these complexes in mesangial cells by global tyrosine and serine/threonine kinome profiling and confirm by siRNA knock-down experiments; and 3) Identify inhibitors of protein kinases that block the specific signaling pathways used by the immune complexes in mesangial cells in vitro and in vivo. We will select small-molecular-mass kinase inhibitors to target the key signaling pathways stimulated by the pathogenic immune complexes and will perform efficacy testing of promising inhibitors using our cultured human mesangial cells model and passive mouse model of IgAN. Relevance: The improved understanding of the pathogenesis of IgAN will identify therapeutic targets for disease-specific therapy of IgAN as well as potential prognostic biomarkers.